Digoxin could improve diaphragm contractility and fatigability if inhibition of sodium-potassium adenosine triphosphatase enhances calcium influx from extracellular sources, or it could impair contractility and worsen fatigue if it impairs maintenance of the membrane potential. We studied the effects of digoxin on isometric force production, fatigue, and recovery in isolated, directly stimulated, guinea pig and rat diaphragms. Digoxin had no effect on maximal twitch or tetanic tensions compared with control diaphragms in either rat (2 ng/ml to 20 micrograms/ml) or guinea pig (2 ng/ml to 2 micrograms/ml) hemidiaphragms. Digoxin worsened high frequency fatigue and impaired recovery from fatigue in guinea pigs (200 ng/ml to 2 micrograms/ml) but not in rat (2 micrograms/ml) hemidiaphragms. We conclude that digoxin has no effect on diaphragm contractility. Hypopolarization of the membrane potential is the likely cause for the increased fatigability. The difference in responsiveness between species is likely due to insensitivity of rat sodium-potassium adenosine triphosphatase to digoxin.